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| − | <h5>Gene Design</h5>
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| − | <p>Chlorite Dismutase is the second enzyme utilized in a number of bacterial species' perchlorate reducing pathway. In their original work, Thorell et al. cloned, sequenced, and functionally expressed the natural coding sequence for Chlorite Dismutase from <i>Ideonella dechloratans.</i> They determined that the gene was capable of converting ClO<sub>2</sub><sup>-</sup> ions to O<sub>2</sub> gas and Cl<sup>-</sup> ions and that its activity was localized to the periplasm, even though its putative signal peptide had not been cleaved. We designed this Cld(-SP) gene to lack this putative signal sequence. The purpose of this modification was so that once translation was complete, the Chlorite Dismutase enzyme would remain in the cytoplasm of our <i>E. coli.</i> chassis. A C-terminal histidine tag was also added to this translational unit in order to purify the enzymes in the event that O<sub>2</sub> production from living cells proved inefficient. Flanking these BsaI sites were XbaI (5’) and the BBa_ std 1 suffix (3’) to facilitate parts creation for the iGEM Registry. We note that the natural coding sequence contained none of the standard BBa_10 restrictions sites, but did contain a single internal BsaI site which we eliminated with a single base silent substitution.
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| − | <h5>Functionality</h5>
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| − | <p>An oxygen production assay was conducted where transformed <i>E. coli</i> was combined with solid Sodium Chlorite. Almost immediately, the formation of a gas, which was inferred to be oxygen, was noted. This indicates the proper functioning of our synthetic G-Block and aligns with the results of Thorell et. al.
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| − | </p>
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| − | <h5>References</h5>
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| − | <p>
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| − | Thorell, H.D, Karlsson, J., Portelius, E.and Nilsson, T. Biochimica et Biophysica Acta 1577 (2002) 445–451
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| − | Samant,S., Gupta, G., Karthikeyan, S., Haq, S., Sambasivam, N.G., and Sukumaran, S. J Ind Microbiol Biotechnol (2014) 41:1435–1442
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| − | </p>
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